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In Vitro Studies of Nitrate Reductase Activity in Cotton Cotyledons

Effects of Dowex 1-Cl and BSA ¹

Roger C. Fites^b

^a Department of Plant Sciences, Texas A&M University, College Station, Texas 77843, Department of Botany, North Carolina State University, Raleigh, North Carolina 27607

Germinating cotton (Gossypium hirsutum L. cv. Deltapine 16) cotyledons developed two peaks of in vitro nitrate reductase activity; the first was stable in vitro and appeared 24 hours after imbibition; and the second, which was extremely labile in vitro, began to develop after the seedlings had emerged and developed chlorophyll. Nitrite reductase activity peaked only after the seedlings had emerged. Dowex 1-Cl (10%, w/v) and bovine serum albumin (3%, w/v) significantly improved the activity of extracted enzyme; greater improvement occurred as expansion of the cotyledons progressed. The major effect of bovine serum albumin on nitrate reductase activity in cotyledon extracts appeared to be that of making the extracted enzyme more active rather than increasing the amount of nitrate reductase extracted or improving the stability of the extracted enzyme.

Attempts to correlate protease activity with the increasingly labile nitrate reductase activity in expanding cotyledons were unsuccessful. Instead, when extracts containing stable nitrate reductase were mixed with extracts containing labile nitrate reductase, the latter was stabilized. The nature of the "protector" in the stable extracts is not known. It is heat-stable, but apparently does not function in vivo since nitrate reductase in germinating cotton seedlings rapidly declines following a peak of activity at 24 hours. We suggest that the protector may function by preventing nitrate reductase from dissociating into inactive subunits.